Neurons in the nervous system undergo retrograde degeneration in neurodegenerative diseases and after acute neurological insults. This grant was awarded previously to characterize an animal model of retrograde degeneration of neurons in the dorsal lateral geniculate nucleus (dLGN) induced by target ablation, and to identify molecular mediators of this cell death. We found that this retrograde neurodegeneration is apoptosis, unequivocally defined by its structure, mediation by Bax (a multidomain Bcl-2 family death effector) and p53, and caspase-3 signaling. This cell death emerges with accumulation of perikaryal mitochondria, oxidative damage to DNA, and subcellular translocations of death effectors, and is modulated by neuronal nitric oxide synthase (nNOS). Previous and new experiments, using in situ cell imaging, show that preapoptotic, target-deprived dLGN neurons accumulate mitochondria prior to cell body shrinkage. We hypothesize that these mitochondria are derived from the axon/synaptic terminals. In this grant renewal we will use our model of apoptosis in mouse brain, in which dLGN neurons undergo apoptosis over 7 days after occipital cortex ablation, to study mitochondrial mechanisms of apoptosis in neurons in vivo. In Aim 1 we will identify sources of the accumulating mitochondria and will test the hypothesis that mitochondria return via dynein motors to the dLGN neuron cell body from the remote site of injury in an altered state defined by their capacity for generating reactive oxygen species (ROS) and content of BH3-only death proteins (Bad, Puma, and Noxa). New experiments also suggest that preapoptotic dLGN neurons accumulate intracellular Ca2+. In Aim 2 we will identify possible mechanisms of intracellular Ca2+ accumulation and the preapoptotic roles of mitochondrial Ca uptake and calcineurin-mediated Bad dephosphorylation and mitochondrial translocation. In Aim 3 we will examine the hypothesis that nNOS activation in target-deprived dLGN neurons leads to peroxynitrite production, intracellular Zn2+ accumulation, and mitochondrial dysfunction. This work can define a new mitochondrial mechanism for target deprivation-induced neurodegeneration and can improve the understanding of the cellular and molecular mechanisms of neuronal apoptosis in vivo [unreadable] [unreadable]